m6A Essentials Antibody Sampler Kit
cat.: HAK21121
Product Type: Antibody Sampler Kit
Species reactivity: Human, Mouse, Rat
Applications: WB, IHC-P
Form: Liquid
Storage condition: Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Storage buffer: 1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Uniprot #: SwissProt: Q86U44 Human | Q8C3P7 Mouse | Q9HCE5 Human | Q3UIK4 Mouse | Q15007 Human | Q9ER69 Mouse | Q9C0B1 Human | Q8BGW1 Mouse | Q2A121 Rat | Q6P6C2 Human | Q3TSG4 Mouse | D3ZKD3 Rat | Q9BYJ9 Human | P59326 Mouse | Q9Y5A9 Human | Q91YT7 Mouse
Entrez Gene: 361035 Rat | 295428 Rat | 499020 Rat | 296467 Rat | 313053 Rat
Images
HAK21121_1.jpg Fig1: Western blot analysis of METTL3 on different lysates with Rabbit anti-METTL3 antibody (HA720002) at 1/1,000 dilution.

Lane 1: HeLa cell lysate (20 µg/Lane)
Lane 2: F9 cell lysate (20 µg/Lane)
Lane 3: PC-12 cell lysate (20 µg/Lane)
Lane 4: Mouse heart tissue lysate (40 µg/Lane)
Lane 5: Rat heart tissue lysate (40 µg/Lane)
Lane 6: Mouse spleen tissue lysate (40 µg/Lane)
Lane 7: Rat spleen tissue lysate (40 µg/Lane)

Predicted band size: 64 kDa
Observed band size: 72 kDa

Exposure time: 25 seconds; ECL: K1801;
4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA720002) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21121_2.jpg Fig2: Immunohistochemical analysis of paraffin-embedded mouse testis tissue with Rabbit anti-METTL3 antibody (HA720002) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA720002) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21121_3.jpg Fig3: Immunocytochemistry analysis of HeLa cells labeling METTL3 with Rabbit anti-METTL3 antibody (HA720002) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-METTL3 antibody (HA720002) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
HAK21121_4.jpg Fig4: METTL14 was immunoprecipitated from 0.2 mg A431 cell lysate with HA722910 at 2 µg/25 µl agarose. Western blot was performed from the immunoprecipitate using HA722910 at 1/5,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: A431 cell lysate (input)
Lane 2: HA722910 IP in A431 cell lysate
Lane 3: Rabbit IgG instead of HA722910 in A431 cell lysate

Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 1 minute; ECL: K1801
HAK21121_5.jpg Fig5: Flow cytometric analysis of A431 cells labeling METTL14.

Cells were fixed and permeabilized. Then stained with the primary antibody (HA722910, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
HAK21121_6.jpg Fig6: Western blot analysis of WTAP on different lysates with Rabbit anti-WTAP antibody (HA500525) at 1/1,000 dilution.

Lane 1: K562 cell lysate (12.5 µg/Lane)
Lane 2: Jurkat cell lysate (10 µg/Lane)
Lane 3: Hela cell lysate (10 µg/Lane)
Lane 4: HepG2 cell lysate (10 µg/Lane)
Lane 5: 293T cell lysate (10 µg/Lane)
Lane 6: NIH/3T3 cell lysate (10 µg/Lane)
Lane 7: PC-12 cell lysate (10 µg/Lane)

Predicted band size: 44 kDa
Observed band size: 55 kDa

Exposure time: 30 seconds;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA500525) at 1/1,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature.
HAK21121_7.jpg Fig7: Immunohistochemical analysis of paraffin-embedded human uterus tissue with Rabbit anti-FTO antibody (ET1705-89) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1705-89) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21121_8.jpg Fig8: Western blot analysis of ALKBH5 on different lysates with Rabbit anti-ALKBH5 antibody (HA721628) at 1/1,000 dilution.

Lane 1: HEK-293 cell lysate (20 µg/Lane)
Lane 2: Jurkat cell lysate (20 µg/Lane)
Lane 3: HeLa cell lysate (20 µg/Lane)
Lane 4: HepG2 cell lysate (20 µg/Lane)
Lane 5: MCF7 cell lysate (20 µg/Lane)
Lane 6: F9 cell lysate (20 µg/Lane)
Lane 7: PC-12 cell lysate (20 µg/Lane)
Lane 8: Rat testis tissue lysate (40 µg/Lane)

Predicted band size: 44 kDa
Observed band size: 35~44 kDa

Exposure time: 30 seconds;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721628) at 1/1,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:100,000 dilution was used for 1 hour at room temperature.
HAK21121_9.jpg Fig9: Immunohistochemical analysis of paraffin-embedded mouse testis tissue with Rabbit anti-ALKBH5 antibody (HA721628) at 1/2,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA721628) at 1/2,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21121_10.jpg Fig10: Western blot analysis of YTHDF1 on different lysates with Rabbit anti-YTHDF1 antibody (HA721302) at 1/1,000 dilution.

Lane 1: HepG2 cell lysate (10 µg/Lane)
Lane 2: Hela cell lysate (10 µg/Lane)
Lane 3: 293T cell lysate (10 µg/Lane)
Lane 4: MCF-7 cell lysate (10 µg/Lane)
Lane 5: NIH/3T3 cell lysate (10 µg/Lane)
Lane 6: PC-12 cell lysate (10 µg/Lane)
Lane 7: HCT116 cell lysate (10 µg/Lane)
Lane 8: Mouse testis liver tissue lysate (20 µg/Lane)
Lane 9: Rat testis tissue lysate (20 µg/Lane)
Lane 10: Mouse brain tissue lysate (20 µg/Lane)
Lane 11: Rat brain tissue lysate (20 µg/Lane)

Predicted band size: 61 kDa
Observed band size: 69 kDa

Exposure time: 5 minutes;

10% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721302) at 1/1,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:300,000 dilution was used for 1 hour at room temperature.
Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.